Capillary electrophoresis (CE) is a microscale separation technique which has a number of practical advantages over conventional separation methods. Among them are high separation efficiency, high speed, and small sample size. However, the small sample size used in CE puts a high demand on detector sensitivity.
A mass spectrometer (MS) is a universal detector. It can provide information regarding molecular masses or the structure of compounds with high sensitivity. The first on-line CE-MS was demonstrated by Smith and co-workers in 1987 (J. A. Olivares et al., Anal Chem. 1987, 59, 1230-1232). In on-line CE-MS, the CE eluate is introduced directly into a mass spectrometer. Analytes are distinguished not only by their migration times but also by their molecular masses or fragmentation pattern. CE-MS is thus very useful in the analysis of small, complex samples, and is especially well-suited for analysis of biological materials.
CE has been coupled with many types of mass spectrometers. The ion utilization efficiency in quadrupole MS for full-scan spectra is low. CE is thus preferably coupled with mass spectrometers having higher ion utilization efficiency such as Fourier transform ion cyclotron resonance mass spectrometers (S. A. Hofstadler et al., J. Am. Chem. Soc. 1993, 115, 6983-6984), ion-trap mass spectrometers (J. Henion et al., Anal. Chem. 1994, 66, 2103-2109), and time-of-flight mass spectrometers (L. Fang et al., Anal. Chem. 1994, 66, 3696-3701). Since TOF-MS has high transmission efficiency and can tolerate relatively high background pressures, it may provide the best sensitivity achievable with the current alternatives. Complete mass spectra can be recorded in a single event. Up to 100000 mass spectra can be produced in a second. The high speed of TOF-MS is suitable for the detection of analytes separated by CE, since the duration of the peaks in CE separation is only several seconds. Also, in principle, an unlimited mass range can be achieved, which is suitable for the study of large biomolecules.
Currently, the major challenge in CE-MS is the development of interfaces for introducing the liquid flow into the mass spectrometer without significant loss in the performance of either CE or MS. CE is typically coupled to MS using electrospray ionization (ESI) (M. Yamshita et al., J. Phys. Chem. 1984, 88, 4451-4459; E. C. Huang et al., Anal. Chem. 1990, 62, 713A-725A; A. T. Blades et al., Anal. Chem. 1991, 63, 2109-2114) or continuos-flow fast-atom bombardment (CE-FAB) (R. M. Caprioli et al., Anal. Chem. 1986, 58, 2949-2954; M. A. Moseley et al., J. Chromatogr. 1990, 516, 167-173; R. M. Caprioli et al., J Chromatogr. 1989, 480, 247-257; N. J. Reinhoud et al., Rapid Commun. Mass Spectrom. 1989, 3, 348-351.). MS generates structural information by ascertaining mass to charge ratios for the various ionic species produced by fragmentation of the analyte of interest. Typically, these ionic species have a single charge due to their association with a proton (H.sup.+). Since electrospray produces multiply-charged molecular species, ESI permits characterization of analytes having molecular weights significantly higher than the upper mass limit of the spectrometer. However, because ESI must be performed at atmospheric pressure, only a fraction of the ions produced is skimmed into the MS, sharply reducing the efficiency of the detection.
Recently, several other interfaces have been developed for coupling separation techniques with MS. These include a pulsed sample introduction interface (A. Wang et al., Anal. Chem. 1992, 64, 769-775; A. Wang et al., Anal. Chem. 1994, 66, 3664-3675), a continuous-flow matrix-assisted laser desorption/ionization (MALDI) interface (L. Li et al., Anal. Chem. 1993, 65, 493-495; D. A. Nagra et al., J. Chromatogr. A 1995, 711, 235-245), and an aerosol MALDI interface (K. K. Murray et al., Anal. Chem. 1993, 63, 2534-2537; X. Fei et al., Anal. Chem. 1996, 68, 1143-1147). These new interfaces show good potential to couple CE with MS; however, a make-up solvent is usually required. A make-up solvent is used to increase the total flow rate into the system, for example, to support droplet formation. Supplying a make-up solvent necessitates introduction of a large amount of electrolyte, diluting the concentration of the analyte. In addition, the added electrolytes contribute to chemical noise. The detection limit of proteins in CE-MS is typically in the high femtomole range. However, the concentration limit of detection is in the range of 10.sup.-6 M. This is not sufficient for the study of many biological samples. In order to improve detection limits, new interfaces with high ionization efficiency and high sample utilization are needed.